In Vivo Photoacoustic Imaging: Brain Research Application Vassiliy Tsytsarev, University of Maryland School of Medicine Most of the presented data have been obtained in: Optical Imaging Laboratory Department of Biomedical Engineering Washington University, St. Louis

What is Photoacoustic? Photoacoustic imaging - a hybrid biomedical imaging modality, is developed based on the photoacoustic effect Photoacoustic effect – discovered by Alexander Bell in 1880; he showed that thin discs emitted sound when exposed to a beam of sunlight that was rapidly interrupted with a rotating slotted disk.Alexander Bell soundbeamsunlight

How it works? Laser Ultrasound Ultrasound trasducer

PA Imaging: three types of scanning PACT image of the cortical vasculature in a living adult intact rat Dark-field AR-PAM OR-PAM (Open Brain) (Wang et al., 2003; Stein et al., 2008)

Wavelength-tunable laser system Condenser lens Pinhole Microscope objective Correction lens Ultrasonic transducer Right-angle prism Silicone oil layer Water tank Scanner Polyethylene membrane Acoustic lens Scheme of optical-resolution photoacoustic microscopy (OR-PAM) Hu et al, 2007; (Wang et al., 2003; Stein et al., 2008)

Intact Rat PA Imaging

Open Brain PA Imaging

Functional Brain Imaging: Transcranial Application (Hu and Wang, 2009) In vivo PACT image of the cerebral vascular response to right-side whisker stimulation (intact rat) The hemodynamic response due to whisker stimulation is shown in blue and red and is superimposed on the cortical vascular image shown in gray.

PA Microscopy: 2-Wavelengths Functional Imaging Hu, Maslov, Tsytsarev and Wang, 2009

Photoacoustic imaging of the vascular response Real-time monitoring of photoacoustic signals at specific excitation wavelengths reveals vascular dynamics, such as changes in blood volume and oxygen saturation, in response to electrical or physiological stimulation or epileptic seizures

Neurovascular coupling: response to electrical stimulation Astrocyte Neuron Electrode Electrical stimulation may cause neurons to release various neurotransmitters (Glutamate [Glu], GABA, ATP and NO). These reactions drive the vessel to either vasoconstriction or vasodilatation GABA Glu K+, NO ATP Blood vessel

Photograph of exposed mouse brain surface with introduced microelectrode Microelectrode Cranial opening 5 mm

Photoacoustic imaging of the brain microvasculature Photograph of the microelectrode Min Max 1.0 SO 2 1mm The oxygen saturation (SO 2 ) mapping is shown as a superposition in color scale. A line-scan monitoring of the vascular response was performed along the dashed yellow line. 400 μm Optical absorption 0.6

Vascular response to electrical stimulation Transverse Axial 10 μm Transverse Axial Stimulation

Vascular response to electrical stimulation

Vasoconstriction and vasodilatation 50 s 130 s 230 s 150 s 250 s 10 s 50 s 126 s 217 s 150 s 280 s 25 µm 50 µm Optical absorption Min Max Electrical stimulations 150 µA100 µA

Time courses of the vessel cross-sectional area under various stimulation intensities Vasoconstriction and vasodilatation are observed, and the response duration is positively correlated with stimulation intensity Stimulation 1 Stimulation 2 Each stimulation consisted of a train of four 0.3 ms pulses at 300 Hz 150 μA 300 μA 350 μA 110 μA 100 μA

Transcranial imaging of the single blood vessel 400 μm Studied vessel Stimulation time (s) medial rostral Transcranial images of stimulation-induced vasodilatation a)Transcranial brain image b)Crossectional monitoring c)Time courses of the electrical- stimulation-induced vessel size (a) (b) (c)

Epileptic Seizures: Single Vessel Monitoring

Summary l Optical-resolution photoacoustic microscopy (OR- PAM) imaged two types of vascular response to electrical stimulation: n Vasoconstriction n Vasodilatation l OR-PAM clearly and reliably imaged the vascular response to electrical stimulation at the capillary level with a temporal resolution of one second. l OR-PAM is a promising tool for in vivo studies of neurovascular coupling under a variety of experimental conditions invasively as well as transcranially

Epileptic seizures accompanied by two vessels vasodilatation (PA Imaging) Vessel 1 Vessel 2 EEG 10 s / 2 mV 25 μm

Conlusions and Perspecives: -Current application: animal experiment -Clinical application: questionable -Perspective: biomarcer hybridozation